Numerous simple visual test devices have been developed for the analysis of body fluids in order to determine component analyte concentrations. These tests include such devices as means for detecting, for example, glucose or other sugars, cholesterol, proteins, ketones, uric acid, phenylalanine, or enzymes in either blood or urine. Yet, it has been particularly difficult to perform visual measurements of these constituents in whole blood. This difficulty lies in the problems associated with visual interference caused by the presence of red blood cells in whole blood. The deep red coloration of red blood cells or free hemoglobin seriously interferes with such whole blood visual analyte concentration analysis.
Means have been proposed for separating and removing highly colored red cells and hemoglobin from whole blood prior to analysis. Some of the simpler methods involve the use of a carrier member impregnated with a test reagent composition and coated with a semipermeable membrane which effectively acts as a means for screening out cells or large molecules such as hemoglobin. This semipermeable membrane permits the passage of smaller molecules or ions in the solution. A substantially clear fluid containing the constituent diffuses into the test reagent in the carrier to cause a chromogenic reaction with the reagent.
Other methods have included taking whole blood samples and placing such samples on a bicomponent reagent strip. After a predetermined time period, the blood sample is blotted to remove excess blood from the top of the strip. At that point, constituents of the whole blood sample migrate onto the strip, and then react with reagent molecules embedded in the reagent strip, and a visual comparison of the resultant color of the reacted blood is made to a chart or standard.
These methods are cumbersome and generally laborious and require at least one extra manipulative step, such as wiping, blotting or rinsing with water. This amounts to considerable loss in time and more importantly, accuracy and efficiency. Moreover, the filtering membrane screens out larger molecules in solution, which precludes these molecules from reaching the test reagent. This sometimes renders these methods inoperative for particularly needed determinations, such as determination of glucose concentration levels. These methods are also technique-dependent and difficult for untrained operators to perform in a reproducible fashion.
Additional methods provide for the drawing of a whole blood sample, then allowing the blood to clot. Once clotted, the blood is centrifuged to separate cellular components from fluid components. These methods require equipment generally found only in specialized settings and typically are more labor intensive than the previously mentioned methods.
Other test systems may comprise a single matrix which contains both a separating component and a test reagent impregnated in the reagent in such a way that the whole blood first contacts the separating component to form a substantially colorless fluid which then contacts the test reagent. In employing such a single matrix test system the separating component and the test reagent must be stable and reactive in each other's presence. The matrix must be designed so that the analyte contained in the whole blood sample reaches the area of the matrix where the response is read by a meter, in a state that is substantially free of any blood coloration. In such an embodiment, a porous support is first coated or impregnated with the test reagent and subsequently the surface of the matrix is coated or impregnated with the separating component. In such a test matrix device, the whole blood contacts first the separating component and the test response is observed in an area not initially in contact with the blood and to which the substantially colorless fluid has migrated.
Examples of such single matrix test strips included separating components which have been found to be, among other things, water-soluble salts, amino acids and carbohydrates such as mannitol. Some of these chemicals cause hemolysis, which is the release of cellular constituents, including hemoglobin. The salts found effective as separating components are non-volatile and do not decompose to any extent under the conditions of preparing and utilizing the test device. The salts have been defined as having solubility in distilled water of at least about 1 gram per liter at 20.degree. C.
Yet, in many instances, fluid containing red blood cells or hemoglobin continues to seep through the matrix despite the presence of the separating component causing the test to mix reagent with colored blood components. When this occurs, accuracy is compromised, and visual comparison is difficult.
It is thus an object of the present invention to provide a unitary test device, wherein during one step the user can apply an unmeasured sample of whole blood and determine the concentration level of an analyte in the whole blood sample.
It is therefore another object of the present invention to provide a unitary test device wherein the test device, whether single or multi-layer, contains separating means as well as test reagent.
It is a further object of the present invention to form a test device consisting of a single matrix wherein whole blood samples can be applied to one side of the matrix and a visual comparison of the analyte concentration level can be made at the opposite side of the test strip. Alternatively, in a longitudinal transport device, such readings can be made on a second portion of the test strip after wicking away from the first portion of the matrix, where the solution sample has been applied.
It is yet a further object of the present invention to determine glucose levels in whole blood samples where a wholly unmeasured sample of whole blood is applied to a single side of a reagent strip. The separating component and test reagent are coated on or trapped within the reagent strip. The test strip then effectively and simultaneously separates the blood into constituent clear fluid and red blood cells and reacts with the glucose in the clear fluid in a manner enabling one to determine, visually, glucose concentration levels of the wholly unmeasured whole blood sample.
It is finally an object of the present invention to provide a test device such that whole blood is analyzed in a single manipulative step for selected molecular constituents such as glucose by a combination of separation means and detection means.